Summary
Pseudomonas is a large and diverse genus of Gammaproteobacteria. To provide a framework for discovery of evolutionary and taxonomic relationships of these bacteria, we compared the genomes of ...type strains of 163 species and 3 additional subspecies of Pseudomonas, including 118 genomes sequenced herein. A maximum likelihood phylogeny of the 166 type strains based on protein sequences of 100 single‐copy orthologous genes revealed thirteen groups of Pseudomonas, composed of two to sixty three species each. Pairwise average nucleotide identities and alignment fractions were calculated for the data set of the 166 type strains and 1224 genomes of Pseudomonas available in public databases. Results revealed that 394 of the 1224 genomes were distinct from any type strain, suggesting that the type strains represent only a fraction of the genomic diversity of the genus. The core genome of Pseudomonas was determined to contain 794 genes conferring primarily housekeeping functions. The results of this study provide a phylogenetic framework for future studies aiming to resolve the classification and phylogenetic relationships, identify new gene functions and phenotypes, and explore the ecological and metabolic potential of the Pseudomonas spp.
Alkyne-containing natural products have been identified from plants, insects, algae, fungi, and bacteria. This class of natural products has been characterized as having a variety of biological ...activities. Polyynes are a subclass of acetylenic natural products that contain conjugated alkynes and are underrepresented in natural product databases due to the fact that they decompose during purification. Here we report a workflow that utilizes alkyne azide cycloaddition (AAC) reactions followed by LC-MS/MS analysis to identify acetylenic natural products. In this report, we demonstrate that alkyne azide cycloaddition reactions with p-bromobenzyl azide result in p-bromobenzyl-substituted triazole products that fragment to a common brominated tropylium ion. We were able to identify a synthetic alkyne spiked into the extract of Anabaena sp. PCC 7120 at a concentration of 10 μg/mL after optimization of MS/MS conditions. We then successfully identified the known natural product fischerellin A in the extract of Fischerella muscicola PCC 9339. Lastly, we identified the recently identified natural products protegenins A and C from Pseudomonas protegens Pf-5 through a combination of genome mining and RuAAC reactions. This is the first report of RuAAC reactions to detect acetylenic natural products. We also compare CuAAC and RuAAC reactions and find that CuAAC reactions produce fewer byproducts compared to RuAAC but is limited to terminal-alkyne-containing compounds. In contrast, RuAAC is capable of identification of both terminal and internal acetylenic natural products, but byproducts need to be eliminated from analysis by creation of an exclusion list. We believe that both CuAAC and RuAAC reactions coupled to LC-MS/MS represent a method for the untargeted identification of acetylenic natural products, but each method has strengths and weaknesses.
Advanced gravitational-wave detectors such as the laser interferometer gravitational-wave observatories (LIGO) require an unprecedented level of isolation from the ground. When in operation, they ...measure motion of less than 10−19 m. Strong teleseismic events like earthquakes disrupt the proper functioning of the detectors, and result in a loss of data. An earthquake early-warning system, as well as a prediction model, have been developed to understand the impact of earthquakes on LIGO. This paper describes a control strategy to use this early-warning system to reduce the LIGO downtime by 30%. It also presents a plan to implement this new earthquake configuration in the LIGO automation system.
Ten strains representing four lineages of the Pseudomonas fluorescens group (P. chlororaphis, P. corrugata, P. koreensis, and P. fluorescens subgroups) were evaluated for toxicity to the tobacco ...hornworm Manduca sexta and the common fruit fly Drosophila melanogaster. The three strains within the P. chlororaphis subgroup exhibited both oral and injectable toxicity to the lepidopteran M. sexta. All three strains have the gene cluster encoding the FitD insect toxin and a ΔfitD mutant of P. protegens strain Pf-5 exhibited diminished oral toxicity compared to the wildtype strain. Only one of the three strains, P. protegens Pf-5, exhibited substantial levels of oral toxicity against the dipteran D. melanogaster. Three strains in the P. fluorescens subgroup, which lack fitD, consistently showed significant levels of injectable toxicity against M. sexta. In contrast, the oral toxicity of these strains against D. melanogaster was variable between experiments, with only one strain, Pseudomonas sp. BG33R, causing significant levels of mortality in repeated experiments. Toxin complex (Tc) gene clusters, which encode insecticidal properties in Photorhabdus luminescens, were identified in the genomes of seven of the ten strains evaluated in this study. Within those seven genomes, six types of Tc gene clusters were identified, distinguished by gene content, organization and genomic location, but no correlation was observed between the presence of Tc genes and insect toxicity of the evaluated strains. Our results demonstrate that members of the P. fluorescens group have the capacity to kill insects by both FitD-dependent and independent mechanisms.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The products synthesized from a hybrid polyketide synthase/nonribosomal peptide synthetase gene cluster in the genome of Pseudomonas fluorescens Pf-5 were identified using a genomics-guided strategy ...involving insertional mutagenesis and subsequent metabolite profiling. Five analogs of rhizoxin, a 16-member macrolide with antifungal, phytotoxic, and antitumor activities, were produced by Pf-5, but not by a mutant with an insertion in the gene cluster. The five rhizoxin analogs, one of which had not been described previously, were differentially toxic to two agriculturally important plant pathogens, Botrytis cinerea and Phytophthora ramorum. The rhizoxin analogs also caused swelling of rice roots, a symptom characteristic of rhizoxin itself, but were less toxic to pea and cucumber roots. Of the rhizoxin analogs produced by Pf-5, the predominant compound, WF-1360 F, and the newly described compound 22Z-WF-1360 F were most toxic against the two plant pathogens and three plant species. These rhizoxin analogs were tested against a panel of human cancer lines, and they exhibited potent but nonselective cytotoxicity. This study highlights the value of the genomic sequence of the soil bacterium P. fluorescens Pf-5 in providing leads for the discovery of novel metabolites with significant biological properties.
The GacS/GacA signal transduction system is a central regulator in Pseudomonas spp., including the biological control strain P. fluorescens Pf-5, in which GacS/GacA controls the production of ...secondary metabolites and exoenzymes that suppress plant pathogens. A whole genome oligonucleotide microarray was developed for Pf-5 and used to assess the global transcriptomic consequences of a gacA mutation in P. fluorescens Pf-5. In cultures at the transition from exponential to stationary growth phase, GacA significantly influenced transcript levels of 635 genes, representing more than 10% of the 6147 annotated genes in the Pf-5 genome. Transcripts of genes involved in the production of hydrogen cyanide, the antibiotic pyoluteorin and the extracellular protease AprA were at a low level in the gacA mutant, whereas those functioning in siderophore production and other aspects of iron homeostasis were significantly higher in the gacA mutant than in wild-type Pf-5. Notable effects of gacA inactivation were also observed in the transcription of genes encoding components of a type VI secretion system and cytochrome c oxidase subunits. Two novel gene clusters expressed under the control of gacA were identified from transcriptome analysis, and we propose global-regulator-based genome mining as an approach to decipher the secondary metabolome of Pseudomonas spp.
To compare work of breathing (WOB) indices between two nCPAP settings and two levels of HFNC in a crossover study.
Infants with a CGA 28-40 weeks, baseline of HFNC 3-5 lpm or nCPAP 5-6 cmH2O and ...fraction of inspired oxygen ≤40% were eligible. WOB was analyzed using respiratory inductive plethysmography (RIP) for each of the four modalities: HFNC 3 and 5 lpm, nCPAP 5 and 6 cmH2O. N=20; Study weight 1516 g (±40 g).
Approximately 12,000 breaths were analyzed indicating a high degree of asynchronous breathing and elevated WOB indices at all four levels of support. Phase angle values (means) (P<0.01): HFNC 3 lpm (114.7°), HFNC 5 lpm (96.7°), nCPAP 5 cmH2O (87.2°), nCPAP 6 cmH2O (80.5°). The mean phase relation of total breath (PhRTB) (means) (P<0.01): HFNC 3 lpm (63.2%), HFNC 5 lpm (55.3%), nCPAP 5 cmH2O (49.3%), nCPAP 6 cmH2O (48.0%). The relative labored breathing index (LBI) (means) (P≤0.001): HFNC 3 lpm (1.39), HFNC 5 lpm (1.31), nCPAP 5 cmH2O (1.29), nCPAP 6 cmH2O (1.26). Eighty-two percent of the study subjects-respiratory mode combinations displayed clustering, in which a proportion of breaths either occurred predominantly out-of-phase (relative asynchrony) or in-phase (relative synchrony).
In this study, WOB indices were statistically different, yet clinically similar in that they were elevated with respect to normal values. These infants with mild-to-moderate respiratory insufficiency demonstrate a meaningful elevation in WOB indices and continue to require non-invasive respiratory support. Patient variability exists with regard to biphasic clustered breathing patterns and the level of supplemental fraction of inspired oxygen ≤40% alone does not provide guidance to the optimal matching of WOB indices and non-invasive respiratory support.
Kangaroo mother care (KMC) is defined as prolonged skin to skin care between a mother and infant with the infant lying in prone position on mom's chest. KMC decreases morbidity and mortality and ...promotes physiologic stability. The aim of this study is to measure work of breathing (WOB) during KMC in very low birth weight (VLBW) infants on non-invasive respiratory support.
A prospective observational pilot study was conducted comparing WOB indices during standard care (SC) and KMC. Respiratory inductive plethysmography (RIP) measured WOB indices non-invasively: phase angle and labored breathing index. VLBW infants who were stable on non-invasive respiratory support were randomized to receive RIP measurements during KMC or during SC first. Summary statistics and mixed linear models were used to compare WOB and vital signs.
A total of 32 infants were consented for the study, data collection and analysis was completed on 28 infants. There were no significant differences in mean phase angle during KMC or SC (73.5±4.6 SE deg vs 66.8±3.9 SE deg, p = 0.25). No differences in WOB and vital signs were detected. Controlling for respiratory support or randomization/first location did not change the results.
In this pilot cohort, infants demonstrated no differences in work of breathing indices or oxygen saturation during KMC or SC while receiving non-invasive respiratory support. KMC appears to be safe and well tolerated with no worsened WOB. Larger studies should be performed to confirm our findings.
Summary
Gene expression profiles of the biological control strain Pseudomonas protegens Pf‐5 inhabiting pea seed surfaces were revealed using a whole‐genome oligonucleotide microarray. We identified ...genes expressed by Pf‐5 under the control of two global regulators (GacA and RpoS) known to influence biological control and secondary metabolism. Transcript levels of 897 genes, including many with unknown functions as well as those for biofilm formation, cyclic diguanylate (c‐di‐GMP) signalling, iron homeostasis and secondary metabolism, were influenced by one or both regulators, providing evidence for expression of these genes by Pf‐5 on seed surfaces. Comparison of the GacA and RpoS transcriptomes defined for Pf‐5 grown on seed versus in broth culture overlapped, but most genes were regulated by GacA or RpoS under only one condition, likely due to differing levels of expression in the two conditions. We quantified secondary metabolites produced by Pf‐5 and gacA and rpoS mutants on seed and in culture, and found that production profiles corresponded generally with biosynthetic gene expression profiles. Future studies evaluating biological control mechanisms can now focus on genes expressed by Pf‐5 on seed surfaces, the habitat where the bacterium interacts with seed‐infecting pathogens to suppress seedling diseases.
Estimating nest success and evaluating factors potentially related to the survival rates of nests are key aspects of many studies of avian populations. A strong interest in nest success has led to a ...rich literature detailing a variety of estimation methods for this vital rate. In recent years, modeling approaches have undergone especially rapid development. Despite these advances, most researchers still employ Mayfield’s ad–hoc method (Mayfield, 1961) or, in some cases, the maximum–likelihood estimator of Johnson (1979) and Bart & Robson (1982). Such methods permit analyses of stratified data but do not allow for more complex and realistic models of nest survival rate that include covariates that vary by individual, nest age, time, etc. and that may be continuous or categorical. Methods that allow researchers to rigorously assess the importance of a variety of biological factors that might affect nest survival rates can now be readily implemented in Program MARK and in SAS’s Proc GENMOD and Proc NLMIXED. Accordingly, use of Mayfield’s estimator without first evaluating the need for more complex models of nest survival rate cannot be justified. With the goal of increasing the use of more flexible methods, we first describe the likelihood used for these models and then consider the question of what the effective sample size is for computation of AICc. Next, we consider the advantages and disadvantages of these different programs in terms of ease of data input and model construction; utility/flexibility of generated estimates and predictions; ease of model selection; and ability to estimate variance components. An example data set is then analyzed using both MARK and SAS to demonstrate implementation of the methods with various models that contain nest–, group– (or block–), and time–specific covariates. Finally, we discuss improvements that would, if they became available, promote a better general understanding of nest survival rates.
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